Summary (Digital Analysis and Development) Over the past 15 years, advances in molecular biology, genetic engineering and microscopy technology have provided biomedical researchers with the capability to use fluorescence microscopy to capture the dynamics of multiple processes occurring in living animals or to characterize the structure and constitution of tissue volumes approaching centimeter scale. The unprecedented size and complexity of this these data has extraordinary potential for enriching our understanding of the mechanisms underlying physiology and disease. However, realizing this potential requires image processing software and hardware powerful enough to address the size and complexity of the data, while being simple enough to be used by mainstream biologists. Since 2002, the IU O'Brien Center has pioneered intravital and 3-dimensional imaging of renal tissues, with the Digital Analysis and Development Core (DAD core) providing consultation, training and software development to facilitate the exploration and quantitative analysis of microscopy data. The ongoing goal of the DAD core is to develop image processing solutions that will help biologists explore and quantify complex image data. During the next grant period, this goal will be addressed via three specific aims: (1) Implement software tools previously developed by the core into an intuitive and efficient software interface. (2) Develop the Distributed and Networked Analysis of Volumetric Image Data (DINAVID) high-performance image analysis system that will support storage, management, visualization, quantitative analysis, and exploration of large (multiple gigabyte) image volumes collected by the 3D tissue cytometry core, and (2) support real-time by remote investigators and (3) Develop new tools for automated image segmentation based upon machine learning.